We propose to study the structures of biological macromolecules and supramolecular assemblies by high resolution electron microscopy and by optical and digital image processing of electron micrographs. Our approach is to extract the maximum possible information from each specimen by using image processing techniques to increase the signal-to-noise ratio of images and to reconstruct specimens in three dimensions in order to sort out details which overlap when three-dimensional structures are imaged by projecting them onto a plane when electron micrographs are recorded. We will combine information from specimens prepared by negative staining, unstained specimens, freeze-drying and shadowing, and freeze-fracture replication. We are presently actively studying two systems: 1) Vesicle crystals of beefheart cytochrome c oxidase and 2) Eukaryotic ribosome tetramers and tetramer crystals. We have already developed many of the techniques of image processing required to effect signal averaging in two-dimensional images, three-dimensional reconstruction from images of tilted specimens and surface reconstruction from images of specimens which have been freeze-dried and shadowed. In order to interpret two-dimensional and three-dimensional structures of these objects, we are using techniques for specifically labeling different functional portions of the structures using both: a) Antibodies--Fab-fragments from conventional antisera and from highly specific monoclonal antibodies and b) Heavy atom labels--undecagold cluster compounds. We present preliminary results in the main portion of this proposal.